Abstract B031: Investigation of the ubiquitin ligase RNF31 as a mediator of immune evasion in advanced prostate cancer
Authors: Tianyi Liu, Haolong Li, Tony Junjie Hua, Hyunjin Shin, Tanushree Shenoy, Martin Sjoestroem, Arian Lundberg, Luke Gilbert, David Quigley, Alan Ashworth, Lewis Lanier, Felix Feng
Journal: Cancer Research (2023)

Abstract Background: Metastatic Castration-Resistance Prostate cancer (mCRPC) is a disease state associated with poor survival. Current FDA-approved therapies are non-curative and can only extend mCRPC patient survival by a few months. Therefore, there is an unmet need to improve therapies for patients with advanced disease. Immunotherapy is an effective treatment strategy for a number of cancers but showed limited success in mCRPC patients. Prostate cancer cells exhibit reduced surface expression of major histocompatibility complex (MHC) molecules as an evasive mechanism from cytotoxic CD8+ T-cells. Natural killer (NK) cells are another class of immune cells with anti-tumor characteristics and can target cancer cells independently of MHC expression. Increased NK cell infiltration and activity in prostate cancer has been associated with favorable patient outcomes, but cancer cells have evolved resistance mechanisms against NK cell targeting. We hypothesize that there are druggable pathways that can be exploited to enhance anti-tumor activities of NK cells and improve efficacy of immunotherapies. Methods: To investigate these pathways, we established and characterized an in vitro co-culture system between prostate cancer and NK cells. We conducted the first targeted pool CRISPR screen identifying genes that, when knockout (KO) in prostate cancer cells, enhance killing by NK cells. We validated the screen by individually KO the gene hits. To explore the potential resistance mechanisms conferred by these hits, we performed RNA seq in control and KO cells. Results: Our screen nominated several known ligands of NK cells including CD58 and PVRL2, as well as RNF31 as the top novel target in promoting NK-mediated kill of cancer cells. RNF31 is upregulated in prostate cancer and is part of a druggable linear ubiquitin chain assembly complex (LUBAC). Single sgRNA validation experiments confirmed that RNF31 KO significantly sensitized LNCaP cells to NK92-mediated depletion. Importantly, RNF31 KO did not affect cancer cell proliferation, suggesting that the observed effects are due to the actions of NK cells. We observed that immune response related genes were upregulated in the RNF31 KO cells compare to the wildtype cells. Furthermore, pharmacologic inhibiting RNF31 via LUBAC inhibitor Hopin-8 sensitizes LNCaP cell to NK cell mediated kill. Conclusions: We anticipate that results from this study will yield insights into a novel tumor-intrinsic mechanism of immune evasion in prostate cancer and evaluate the therapeutic potential of targeting this pathway. We hope this work will help identify new therapeutic options or combination treatments that can improve the efficacy of immunotherapy in prostate cancer and significantly extend the lives of patients with advanced prostate cancer. Acknowledgements: This study was supported by the DOD Early Investigator Research Award to TL. Citation Format: Tianyi Liu, Haolong Li, Tony Junjie Hua, Hyunjin Shin, Tanushree Shenoy, Martin Sjoestroem, Arian Lundberg, Luke Gilbert, David Quigley, Alan Ashworth, Lewis Lanier, Felix Feng. Investigation of the ubiquitin ligase RNF31 as a mediator of immune evasion in advanced prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B031.